Positron Emission Tomography Imaging of Thrombus. The goal of this proposal is to develop a fibrin-specific positron emission tomography (PET) probe for direct imaging of thrombus. Thrombus is implicated in a range of pathologies, e.g. ischemic stroke, myocardial infarction, pulmonary embolism, and deep vein thrombosis. The presence of thrombus changes treatment decisions and imaging is widely used rule in, or out, the presence of thrombus. Despite the different imaging methods available, there is still a strong unmet medical need for direct identification of thrombus. For instance, identification of the source of the embolus in ischemic stroke is critical to patient management, but currently 30- 40% of strokes are of unknown origin. This proposed PET probe may enable identification of stroke source and identify patients who would benefit from more aggressive antithrombotic therapy to prevent secondary stroke. Stroke is the third leading cause of death and the leading cause of morbidity in the US. According to the American Stroke Association, Americans will pay almost $69 billion dollars this year in stroke related medical costs and disability. An imaging tool to identify those at risk for secondary stroke may be expected to positively impact morbidity and mortality associated with stroke. The thrombus probe is based on a peptide that binds to and has specificity for fibrin over circulating fibrinogen or other plasma proteins. Fibrin is present in acute and sub-chronic venous and arterial thrombi in high concentrations, but fibrin is not present in circulating blood. A selective fibrin imaging probe is expected to provide high sensitivity and specificity for thrombus identification. While PET offers molecular specificity, it suffers from low spatial resolution and lacks anatomical landmarks. Hybrid PET - computed tomography (CT) or PET - magnetic resonance imaging (MRI) devices are now available that enable co-registration of the molecular PET image to the vascular anatomical images obtained by CT or MRI. This proposal builds on positive preliminary PET-MRI thrombus imaging data with a prototype PET probe. In Aim 1 we focus on synthesizing a series of fluorine-18 and copper-64 labeled peptides to optimize the imaging properties of thrombus-targeted PET probe. In Aim 2 we will characterize these new probes with respect to their affinity to the fibrin target, their specificity for fibrin over other plasma proteins, and their metabolic stability in the presence of plasma or liver homogenate. In Aim 3 we explore the efficacy of these probes in fresh and aged thrombus models in rats and rabbits using PET-CT and PET-MRI. Dosimetry estimates will be performed on optimized probes and the most promising will be evaluated for acute toxicity. The output of this research will be a novel thrombus imaging probe that may be subsequently developed for human use.